If Kr is acting in the control of NB fate as in the Hb/Kr/Pdm/Cas cascade, Kr mutant NB clones should simply skip the Kr-dependent VA7l fate, resulting in the loss
of a single progeny neuron. In other words, VA7l-lacking Kr mutant NB clones should not carry any ectopic VA2 neurons, as observed in mutant Volasertib order GMC clones. In support of this scenario, we confirmed that Kr mutant NB clones contain one lone VA2 adPN through visualizing specific adPN types using a sparse GAL4 driver ( Pfeiffer et al., 2008) ( Figure 4A). This is very different from the chinmo mutant NB clones in which loss of Chinmo-dependent adPNs was accompanied by an equivalent increase in the cell count of the next Chinmo-independent adPN type, leaving the lineage length unchanged MAPK Inhibitor Library solubility dmso ( Figure 4B). These observations indicate that Kr governs temporal fate transitions in the NB, whereas Chinmo acts in the offspring to refine neuronal temporal identity. We next examined how ectopic Chinmo or Kr might affect adPN development to assess their role as master genes for specifying temporal fate. Such gain-of-function experiments provide clues of their endogenous expression pattern, which is challenging to visualize in real time. In chinmo mutant NB
clones generated in first-instar larvae, expression of transgenic chinmo during neurogenesis effectively restored all the missing glomerular targets ( Figures S3C and S3D). Analogous induction also fully rescued chinmo mutant GMC clones ( Figure S3B). No gain-of-function
phenotype was observed, as ectopic Chinmo failed to elicit any late-to-early Terminal deoxynucleotidyl transferase temporal fate changes in wild-type clones, even among those that normally acquire the D fate, the default fate for all the neurons born within the second Chinmo-required window ( Figures S3E and S3F). Use of various chinmo transgenes, including those expressed uniformly due to lack of the endogenous 5′ UTR, yielded identical outcomes ( Zhu et al., 2006). These results suggest that Chinmo promotes neuron diversity through collaborating with other temporal factors governed by NB temporal identity. By contrast, a transient induction of transgenic Kr severely perturbed adPN development. Single-cell clones, as well as the drastically reduced NB clones, no longer targeted dendrites to specific glomeruli; and their axons barely reached the LH (data not shown). Such rudimentary morphologies prevented any meaningful assessment of neuron types or temporal identity. To determine whether ectopic Kr can specify additional VA7l adPNs may require more sophisticated control over when, where, and at what level the Kr transgene should be induced.